A molecular-genetic approach is required to fully understand the complex developmental and physiological processes that occur within a mammalian organism. The mutation at the mouse pygmy locus leads to a disruption of the overall pattern of growth and development and gives rise to mice that are miniature in size. The aim of this proposal is to continue and expand the studies on the pygmy locus which has been cloned via insertional mutagenesis in transgenic mice. The experimental approach will be to further characterize this mutation at the phenotypic and molecular level. A detailed histological analysis of the hypothalamus, pituitary, thyroid and adrenal will be carried out since this axis seems to be perturbed in the mutants. The hormone levels secreted by these tissues and those involved in growth will also be thoroughly investigated. The initial aim of the molecular biology experiments will be to isolate the pygmy gene product. This will be achieved by cloning the normal locus and isolating conserved single copy sequences between different species. These sequences will then by hybridized with RNa isolated from embryos at various developmental stages and different adult tissues. A cDNA library will be constructed form the appropriate expressing tissue and the whole coding region isolated, sequenced and used to investigate the developmental and tissue specific expression of the gene. The protein will be overexpressed in E. coli and antibodies generated to elucidate the cellular localization of the protein. Gene therapy and gene disruption experiments will prove unequivocally if the isolated gene is actually responsible for the dwarf phenotype. Finally, the homologous gene from humans will be isolated and used to see if there s a mutation in families suffering from non growth hormone- deficient dwarf syndromes. The eventual aim of these studies is to attempt to understand the physiological role of the pygmy gene product in growth and development. In addition, these investigations will determine the utility of the mutant mice as molecularly authentic murine analogs of non growth hormone- deficient human dwarf syndromes.